Abstract:To identify the fish eggs which were obtained from Enteromorpha algae of Rudong sea area, DNA barcoding technique was used. Our result showed there were no variable sites in mtDNA COI gene sequences among our fish eggs, larvae, adults and Japanese halfbeak (Hyporhamphus sajori), and the genetic distance among them was 0. However, there are high variable sites among the eggs, larvae, adults and the other Beloniformes fishes, and the genetic distance among them is 21.9%-26.4%. The Neighbor-joining (NJ) phylogenetic tree also indicated that the eggs, larvae, adults and the Japanese halfbeak formed a monophyletic group. Therefore, it is suggested that the fish eggs which we obtained from Enteromorpha algae of Rudong sea area were Hyporhamphus sajori. Based on our identification result, we further discussed the reasons of Japanese halfbeak spawning in algae and how to use this phenomenon to recover the offshore fish resources.

Abstract:To understand the potential function of pluripotency-associated gene Oct4 in blunt-snout bream (Megalobrama amblycephala), the present study investigated the prokaryotic expression of the recombinant Oct4 protein and generated the rabbit anti-Oct4 polyclonal antibody. Firstly, the expression vector pET32a-MaOct4 was constructed by inserting the C-terminal coding region of Ma-Oct4 into pET32a. Then the expression vector was transformed into Escherichia coli BL21(DE3)pLysS, and the recombinant Oct4 protein was induced by IPTG. After optimization of expression conditions, the protein was largely induced and purified to immunize the New Zealand rabbits. Subsequently, the titer and specificity of the generated antibody were assayed by ELISA and Western blot. The recombinant Oct4 protein was highly induced by 0.5 mmol/L IPTG for 4 h at 37℃. The polyclonal anti-Oct4 antibody effectively recognized the purified recombinant Ma-Oct4 antigen, the induced Ma-Oct4 protein in E.coli, the endogenous Ma-Oct4 protein from the fish embryos, and the overexpressed Ma-Oct4:DsRed fusion protein in HepG2 cells. In conclusion, this research provided an effective antibody to study the potential function of Oct4 in blunt-snout bream.

Abstract:The lateral line system originates from the embryonic lateral line placode. The specification of lateral line placode requires the collaboration of a variety of signalings. In order to study the role of Wnt signaling in the development of the lateral line system in zebrafish, we modulated the Wnt signaling by heating shock of the transgenic zebrafish, hsp:Wnt8a-egfp and hsp:dkk1-egfp, and evaluated the hair cell number by YO-PRO-1 staining. We found that hair cells in the zebrafish lateral line system cannot form normally after overexpression of Dkk1. Furthermore, we found Wnt signaling was also necessary for precursor cells formation using alkaline phosphatase staining and in situ hybridization of Eya1 and Six2b. In summary, Wnt signaling affects the development of lateral line system through regulating lateral line placode formation.

Abstract:To explore a simple and feasible method for drug injection into zebrafish gonad,the integrated surface projection of the sex gonads was firstly figured out by dissection, precise measurements and calculation of ten adult zebrafish. Injections of the Alcian blue dye were tried to determine the in vitro injection path and injection area. Then dextran texas red and sox9a-myc overexpressed plasmid gene with liposome transfection reagent mixture were injected into the gonads. 48 h later,dextran texas red distribution was examined under fluorescence microscopy and Sox9a-myc expression was determined by using Western blot. Based on the relative size and location of the sex gonads, injection site was determined upward about 0.1 cm from the intersect between pectoral fin upper horizontal and the front end vertical line of ventral fin. 48 h after injection, Alcian blue and dextran texas red fluorescence were observed in the gonad and surrounding organ. Meanwhile overexpression of Sox9a-myc was detected in ovary using Western blotting analysis. These results suggest that our established gonadal injection method is effective and feasible, which could facilitate the in vivo study of zebrafish sex gene regulation via gonadal drug administration.

Abstract:In this study, the effect of ocean acidification on sex differentation of marine medaka was studied. Fertilized eggs were incubated at three concentrations of CO₂ (480×10^-6, 1 000×10^-6 and 2 000×10^-6) and after hatching, the fry were bred under control condition (480×10^-6) until adulthood. Phenotypic sex was judged from secondary sexual characteristics. The ratio of male to female in two treatment groups was significantly higher than that in the control group (P<0.01). Genotypic sex was determined by the presence or absence of the DMY gene. In the control group, phenotypic sex completely agreed with genotypic sex. However, in two treatment groups, there are some phenotypical males but genotypic female, which is sex-reversed XX male. The sex reversal rate was 38.4% and 47.5% at 1 000×10^-6 and 2 000×10^-6 concentration of CO₂, respectively.

Abstract:Four isonitrogenous and isoenergetic diets were formulated to evaluate the feasibility of partial replacement of fish meal by two plant protein mixtures (composed of dehulled soybean meal and corn gluten meal or rice protein meal, respectively) in 45% fish meal diet and 30% fish meal diets for largemouth bass (Micropterus salmoides). Fish[initial body weight (58.69±0.07) g] were fed over 53 days. The results suggested that there were no significant differences in survival rate among all treatments (P>0.05). 30% fish meal diets-fed fish showed higher final body weight, specific growth rate and feed intake than 45% fish meal diets-fed fish (P<0.05), while feed efficiency ratio, protein efficiency ratio, protein digestibility and total amino acid digestibility showed no significant differences between 30% fish meal diets-fed fish and 45% fish meal diets-fed fish (P>0.05). The fish fed corn gluten meal diets had a significantly higher feed efficiency ratio and protein efficiency rate than fish fed rice protein meal diets (P<0.05), however, feed intake with fish fed corn gluten meal diets was significantly lower than fish fed rice protein meal diets (P<0.05). There were no significant differences in concentrations of crude protein, crude lipid, moisture and ash of the whole body and muscle among all the treatments (P>0.05). Fish fed 30% fish meal diets had significantly higher serum lysozyme activity and serum protein content than fish fed 45% fish meal diets (P<0.05), but no significances occurred between corn gluten meal diets-fed fish and rice protein meal diets-fed fish (P>0.05). No significant differences in red blood cell count and hematocrit among all the treatments were observed (P>0.05). It can be concluded from this study that in the diet for largemouth bass fish meal inclusion level can be replaced from 45% to 30% with the mixtures composed of dehulled soybean meal and corn gluten meal or rice protein meal. The mixture containing corn gluten meal has a better substitution effect than the mixture containing rice protein meal.

Abstract:With the aim to investigate the changes of steroid hormones in the serum and gonadal tissue of Japanese eel(Anguilla japonica) during artificial induction of maturation,we have developed and validated a UPLC-MS/MS method to determine 6 steroids in serum samples simultaneously, including testosterone(T),estrone(E₁),17α-hydroxyprogesterone(17α-OHP),17α, 20β-dihydroxy-4-pregnen-3-one(DHP),estradiol-17β(E₂),estriol(E₃),and 4 steroids (T, DHP, E₂, E₃) in gonadal tissue. The results showed that the average recoveries in serum and gonadal tissue detection were 86.9%-98% and 75.6%-88.1%,respectively; and the detection limit(S/N>10) were 0.1ng/g and 0.1 ng/mL. The steroid hormones in serum and gonad tissue of experimental groups were increased through the hormone injection. Except for the content of E₃, all the other five steroid hormones reached their highest content during spawning, however,the E₂ and DHP in gonadal tissue of experimental groups reached the highest content during dystocia. Conversely, not any of E₁and E₃ in serum, as well as E₂ and E₃ in gonadal tissue were detected through the period in the control group. These results provide preliminary understanding of sex steroid hormones changes during the maturation of Anguilla japonica,moreover,it established a sensitive and stable method to determine the sex steroid hormones in aquatic animals.

Abstract:Studies were conducted on the growth characteristics of one-year-old Alosa sapidissima in pond culture. The cultivation experiment was carried out in No.10 east and No.10 west ponds. The initial culture density was 15 600 ind/hm², and the survival rate was 46.65% in No.10 east pond. By contrast, in No.10 west pond, the initial culture density was 9 600 ind/hm², and the surviving rate was 42.70%. The growth characteristics of Alosa sapidissima in culture cycle were analyzed according to the merged date of growth. The results showed that body length growth increased by 48.3% and body weight growth increased by 286.8% in the condition of 16.4-30.8℃ after 185 days rearing. The relationship between body weight and body length was W=0.0067 L^3.2603 (R²=0.941 6). The power was near 3 which indicated the growth of weight was the same as that of body length. The trend of the daily length increase looked U-shaped while the increase of daily weight looked W-shaped during the experiment. The fatness varied from 1.34 to 1.63, and the highest fatness appeared in the time frame of the highest average water temperature during the summer. The suitable water temperature was 16-30℃, and the optimum water temperature was 19-24℃.

Abstract:The effects of salinity (8,14,20,26,32), temperature (10,15,20℃) and body weight (6 groups) on oxygen consumption rate and ammonia excretion rate of Pleuronectes yokohama juvenile were studied by means of closed breathing experimental method in this study. The results showed that when salinity ranged between 8 and 32, the oxygen consumption rate and ammonia excretion rate were significantly decreased with salinity decline. While temperature was between 10 and 20℃, the oxygen consumption rate and ammonia excretion rate were significantly increased with the increase of temperature. In addition, the oxygen consumption rate and ammonia excretion rate decreased with the increase in weight. The O/N ratio decreased with the increase of salinity and the decrease of temperature, and it showed the O/N ratio ranged from 44.441 to 12.278 and 13.446 to 17.631 respectively. The average Q₁₀ values at these temperature were 1.545 and 2.018 respectively, while no significant differences were found in O/N ratio among different body weight. The energy sources of P. yokohama under the experiment conditions were mostly fat and protein with different ratios, only in the group of salinity 8 the energy sources were fat and sugar.

Abstract:In order to develop Onchidium struma's artificial aquaculture technology and meet the growing market demand, it is crucial to know O. struma's spawning law very well. This study investigated the spawning characteristics within three years and comparatively studied the relationship between spawning and astronomical tides. During three years, O. struma in the same regions was obtained from July to September from 2013 to 2015, and the spawning was recorded each time. After that, the spawning and tidal height was comparatively analyzed. The results showed that O. struma spawned mostly at the neap tide (7^th, 23^rd) every month, and this spawning could last 6-7 days by lunar calendar. The spawning stopped at the advent of 1-2 days before the next spring tide, while the tide would take O. struma larva hatched into the ocean. After about 15 days' development in the sea, O. struma developed into the big veliger and then came back to the beach along with the next tide. It can be concluded that:(1) O. struma spawning rules were related to the tide which showed a peak spawn within the neap tide and reached peak 2-3 days after spawn. (2) Theoretically, it takes 14 d 19 h 55 min from one neap to another neap theoretically. However, in fact, O. struma's spawning cycle was 14 d 21 h 20 min because of the tidal hysteresis.

Abstract:In order to research the change pattern of phytoplankton community structure in pond industrial eco-aquaculture system and explain its ecological mechanism, the experiment was carried out using Megalobrama amblycephala as aquaculture species and setting two stocking density. Shannon-Wiener diversity index and Pielou uniformity index were used to study the phytoplankton community structure in aquaculture area (including Low stocking density aquaculture area and High stocking density aquaculture area), Inlet area, Outlet area, Purification area, Recirculation area, Back area of pond industrial eco-aquaculture system. The results showed that 5 phyla including 92 species have been found, including Chlorophyta 46 species, Cyanophyta 15 species, Euglenophyta 15 species, Bacillariophyta 10 species, Cryptophyta 5 species, Pyrrophyta 1 species. The numbers of phytoplankton species, biodiversity indexes, uniformity indexes, chlorophyta biomass and its proportion accounting for the total phytoplankton followed the order of Inlet area, Purification area, Recirculation area, Back area>Low stocking density aquaculture area >High stocking density aquaculture area>Outlet area. The total biomass of phytoplankton, Cyanophyta and its proportion accounting for the total phytoplankton followed the order of Inlet area, Purification area, Recirculation area, Back area< Low stocking density aquaculture area

Abstract:The seasonal investigation on the functional groups of macroinvertebrates and nutrient status was carried out at 9 sites in the downstream of the East Tiaoxi River from July 2013 to April 2014. The feeding evenness index (j_FD) was used to assess the ecosystem health of the East Tiaoxi River. The results showed that the gather-collectors was the most dominant group among the functional groups of macroinvertebrates in this reach. And the functional groups of macroinvertebrates had no significant difference in four seasons (P>0.05). It suggested that the small organic matters were rich in this reach. The redundancy analysis (RDA) showed that the functional feeding groups were closely related to the total nitrogen in four seasons. The j_FD value indicated that the overall ecosystem health in the downstream of the East Tiaoxi River was relatively good, but the health of some samples was poor. It suggested that there might be point source pollution in this river reach, and the water habitat might be fragmented.

Abstract:The pollutant removal effect towards different cultivation species or quantity on the rotation model by water spinach (Ipomoea aquatica) and cress (Oenanthe stolonifera) has been tested. Contents of the main water quality indexes including total organic carbon(TOC), chemical oxygen demand (COD), chlorophyll (Chl), total nitrogen (TN), ammonium nitrogen (NH₄⁺-N), nitrite nitrogen (NO₂^--N), nitrate nitrogen (NO₃^--N), total phosphorus (TP) and PO₄^3--P were measured. TOC, TN and TP in the sediments have also been tested. The results showed that TOC, Chl, TN, NH₄⁺-N, NO₂^--N, NO₃^--N and PO₄^3--P significantly decreased in Ganlou herring (Mylopharyngodon piceus) pond before the rotation model (only planting water spinach), while TOC, COD, Chl, NH₄⁺-N, NO₂^--N, NO₃^--N, TP significantly decreased in Suzhou economic parental fish pond. TOC, NO₃^--N, TP significantly decreased in Ganlou herring (Mylopharyngodon piceus) pond after the rotation model (only planting cress), while TOC, COD, NH₄⁺-N, NO₃^--N, TP significantly decreased in Suzhou economic parental fish pond. To conclude, TOC, NH₄⁺-N, NO₃^--N, TP in the water and TOC, TN, TP in the sediment have been significantly reduced by this rotation model "water spinach-cress".

Abstract:In order to investigate the water quality and primary productivity of national aquatic germplasm resources conservation area of asian clam and erythroculter ilishaeformis in Dianshan Lake, ten sampling points were chosed in the conservation area in 2015. The spatiotemporal variations of nutrients, chlorophyll-a (Chl.a) and primary productivity, and the relationship between Chl.a and environmental factors were analyzed by comprehensive nutrition status assessment and correlation analysis. The results showed that the concentrations of nutrients, COD_Mn and Chl.a had a significant difference. The results in spring and summer were higher than autumn and winter. But there was no significant difference in the test area compared with the core area. The N/P (nitrogen and phosphorus) ratios were between 5.30-64.66 and they are suitable for the algal growth. The comprehensive nutrition state index in conservation area was between 46.67 to 60.70. It showed the water columns in conservation area were in mild eutrophication state. The variations of Chl.a concentration and primary productivity were between 6.06 to 29.37 mg/m³ and 65.18 to 1 423.11 mg/(m³·d), respectively. The spatiotemporal variations of them were similar. There was a significant difference between the seasons, but there was no significant difference in the test area compared with the core area. The Pearson Correlation Analysis showed there was a significant correlation between the concentrations of Chl.a and TN, TP. The correlation index (R²)of them were 0.501 and 0.432, respectively. The concentrations of nitrogen and phosphorus were higher. They could lead to the water-bloom occurrence in this conservation area.

Abstract:From March 2013 to February 2014,the gill net and cages were conducted on the fish resources in 8 sampling sites of Lake Taihu, aiming to understand the characteristics of Acheilognathinae community structure and diversity in the lake. A total of 50 fish species were collected, belonging to 23 families,and 8 orders, among which, Acheilognathinae,Gobioninae and Engrauulidae had the larger species number and had an obvious tendency of fish resources becoming smaller-scale; as compared with historical data, 57 fish species were not collected,the fish species number and composition of dominant species changed greatly. A total of 7 Acheilognathinae species were collected, belonging to 2 families, the Acheilognathus chankaensis and Acheilognathus macropterus were the main species and allometric growth,other fish were isometric growth.The cluster analysis showed the significant differences in spatial and temporal aspects among the Acheilognathinae in Lake Taihu, clustered into two groups according to the time,from May to December for a group, and the index of N%、W% and IRI showed dominant species was Acheilognathus chankaensis,other time was Acheilognathus macropterus; clustered into two groups according to the space,the Manshan and Gonghu for a group and dominant species was Acheilognathus chankaensis, and other station was Acheilognathus chankaensis.It showed that the diversity index was below the average(1.5-3.5).They were closely related between spatial and temporal variations of fish and distribution patterns of nutrients, environmental factors, plankton, and aquatic plants in the lake.

Abstract:The Yangtze River Estuary is one of the finless porpoise (Neophocaena asiaeorientalis) habitat with the high density. The Dongfengxisha waters located on the southwest side of Chongming Island in the Yangtze Estuary is a distribution hotspot of the Yangtze finless porpoise (N. asiaeorientalis). In order to clarify population dynamics of the finless porpoise in the Dongfengxisha waters, the visual survey and perennial observations were performed in this waters from 2014 to 2016. A total of six investigations covered 224 km length of the line transect and found 68 porpoises in 11 groups. The encounter rate is averagely 0.37 int./km from 0.12 int./km to 1.14 int./km. During 13 months of perennial observations, 340-353 porpoises in 123 groups were witnessed. The average group encounter rate was 0.38 and the average individual encounter rate was 1.07. Visible coefficient method estimated that the finless porpoise population was 11.1-62.7, with the average number of 31.6. Given that the shoal is located in the north of the Dongfengxisha waters and is hard for finless porpoise to survive, the population was corrected as 26.8. Since the finless porpoise ranges throughout the year in the waters, it is necessary to take intensive care and even establish a natural reserve.

Abstract:The investigation was conducted at four sections of Nanji Islands in February and August 2010. One hundred and twenty five species were indentified, including 72 species of molluscs, 23 of macroalgaes, 13 of carapaces, 10 of annelids, 4 of echinoderms and 3 others. 72 species of macrobenthos were collected in winter and 93 species in summer, the average biomass and inhabit density were 3 941.22±2 870.13 g/m², 1 275.44±749.37ind./m² and 4 521.69±3 473.66 g/m², 2 099.07±1 074.82 ind./m², respectively. The dominant species in different seasons have no significant change. The index of Shannon-Wiener's biodiversity was from 1.62 to 2.95, the average being 2.50±0.46. The Margalef's species richness was from 1.99 to 5.88, the average being 4.81±1.31. The Pielou's evenness was from 0.60 to 0.77, the average being 0.71±0.06. The Simpson's dominance was from 0.71 to 0.92, the average being 0.86±0.08. Upon analysis of the macrobenthos community structure by ABC curve, hierarchical clustering analysis and MDS diagrams, the similarity index of community structure between three rocky sections in different seasons were all higher than 50%, and lower than 10% between the only muddy section in winter and summer. The section in Guoxing'ao has the lowest community structural stability, seriously affected by the activities of fishermen.

Abstract:Our previous work suggests that Lb.kefiranofaciens is the dominant bacterial species in Tibetan kefir grains and comprises approximately 95% of all lactic acid bacteria. In this research, bacterial isolation and purification, 16S rRNA gene cloning and sequencing, denaturing gradient gel electrophoresis(DGGE), colony PCR and scanning electron microscope observation were performed to understand non-dominant lactic acid bacteria in kefir grains. Scanning electron microscope observation revealed that the cells of Lb. kefiri are short rod (0.8-1.5×0.6-0.7 μm), which were shorter than Lb. kefiranofaciens.Lb. kefiri cells tended to form aggregates in pure culture condition. After sequencing analysis, 1 of 25 clones was related to Lb. kefiri. For 22 colonies isolated from kefir grains, 5 colonies are Lb. kefiri. In addition, DGGE results showed that Lb. kefiri exists in all kefir grain samples. These results demonstrate that Lb. kefiri exists in kefir grain and remains stable and is non-dominant.

Abstract:In 2012 for the first time we had a 23-day continuous on-site buoy observation at the Nordic seas.Based on the observation data of sea surface air temperature, humidity, wind speed and skin temperature, the turbulence heat flux at air-sea interface was calculated by COARE 3.5 bulk algorithm. The sensible heat flux was 9.34 W/m², the latent heat flux was 31.55 W/m² (ocean to atmosphere as positive).ERA-Interim and OAFlux were international widely used oceanographic and meteorological data sets, then the buoy observation data were compared and verified with two data sets.The results show that the summer surface air-sea temperature difference in the warm area of the Nordic Seas plays a major role in the heat transfer at the air-sea interface,while in the case of low wind speed (<10m/s) in latent heat transfer the leading role is the difference between the humidity between sea and air.ERA-I and OAFlux were slightly overestimated and underestimated by 12.58% and 26.33% respectively for sensible heat fluxes in the summer warm current region of Nordic Sea. The main factors contributing to the bias are the differences between the two data sets and the buoy data for the prediction of the air temperature and the skin temperature.ERA-Interim and OAFlux have a 17.54% and 7.76% over estimate of latent heat flux respectively.The difference comes from the estimation of air & sea humidity.